Recent evidence suggests that polyol pathway-linked dysfunction in galactosemic and diabetic rats is mediated by a hypoxia-like increase in cytosolic free NADH/NAD+ which is due, in diabetic rats, to increased oxidation of sorbitol to fructose (coupled to reduction of NAD+ to NADH) by sorbitol dehydrogenase. These studies were undertaken to test the prediction that since galactitol is a poor substrate for sorbitol dehydrogenase, inhibition of sorbitol dehydrogenase should prevent vascular dysfunction in diabetic, but not in galactosemic rats. Method: Retinal plasma 125 albumin permeation was assessed in galactosemic and streptozotocin-diabetic rats either untreated or given CP-166,572 (an inhibitor of sorbitol dehydrogenase) 200 mg/kg bwt/day or zopolrestat (an aldose reductase inhibitor) 100 mg/kg bwt/day for 6 weeks, and in control rats. Retinal sorbitol and galactitol were assessed by GCMS. Results: Albumin permeation was increased (p<0.0001) in galactosemic and diabetic rats versus controls ((130+20sd ul plasma/g wet wt/min, 137+16, and 75+12, respectively). Zopolrestat decreased polyol levels by 60% in galactosemic and diabetic rats and normalized albumin permeation both groups. CP-166,572, as expected, increased retinal sorbitol levels -4 fold in diabetics from 1409+412 nmol/g wet wt to 5380+1702 (p<0.0001) and normalized albumin permeation (85+24); it had no effect on albumin permeation (143+32) or galactitol levels in galactosemic rats. Conclusion: Sorbitol pathway-linked retinal vascular dysfunction is diabetic rats is mediated by increased oxidation of sorbitol to fructose independent of osmotic effects of high sorbitol levels; different mechanism(s) mediate aldose reductase-linked vascular dysfunction in galactosemic rats. Supported by NIH grant EY06600 the Kilo Foundation.